A Chinese medicinal extract for preventing and treating kiwifruit canker, and its preparation method and application

Abstract

The present invention discloses a traditional Chinese medicine extract for preventing and treating kiwifruit canker, as well as its preparation method and application. The extract comprises the following components: an extract of Radix Isatidis and / or an extract of Rhizoma Cibotii. The extract has the potential to prevent and treat kiwifruit canker pathogens and has the potential to be developed into a new botanical fungicide. Furthermore, the extract is made from traditional Chinese medicinal materials and has less impact on humans, animals, and the environment than chemical pesticides.

Description

Technical Field

[0001] The invention belongs to the technical field of pesticides, and particularly relates to a traditional Chinese medicine extract for preventing and treating kiwifruit canker, and a preparation method and application thereof. Background Art

[0002] Kiwifruit canker is a bacterial disease caused by Pseudomonas syringae pv. actinidiae (Psa), primarily affecting the trunk, branches, new shoots, leaves, and flower buds of kiwifruit. Typical symptoms include polygonal brown spots on leaves surrounded by yellow halos, often causing them to curl. Scorch occurs on young shoots, and flower buds turn brown, failing to open and withering. A milky white mucus oozes from wounds, lenticels, eyes, and leaf scars on branches, which oxidizes to a reddish-brown color. The affected tissue sinks, causing ulcers and rot, and the branches and vines to die. Kiwifruit cultivation in many countries is currently affected by canker.

[0003] Chemical control is the primary prevention and control measure for kiwifruit canker. However, the long-term use of these chemicals has led to drug resistance, environmental pollution, and food safety issues, seriously impacting the green and sustainable development of the kiwifruit industry. Therefore, exploring green control technologies for kiwifruit canker using traditional Chinese medicine resources is a key breakthrough in disease prevention and control. Summary of the Invention

[0004] To address the issues of drug resistance, environmental pollution, and food safety associated with chemical agents used to control kiwifruit canker, the present invention proposes a botanical pesticide. The present invention provides a traditional Chinese medicine extract for controlling kiwifruit canker, as well as its preparation method and application. The extract can achieve the purpose of controlling the kiwifruit canker pathogen by affecting multiple factors, such as cell wall morphology and DNA content. Furthermore, the extract is made from traditional Chinese medicinal materials, which have less impact on humans, animals, and the environment than chemical pesticides.

[0005] In order to achieve the above object, the present invention adopts the following technical solutions:

[0006] The invention provides a traditional Chinese medicine extract for preventing and treating kiwifruit canker, which comprises the following components: a Radix Isatidis extract and / or a Pseudo-Nine-joints extract.

[0007] In the present invention, the Chinese medicinal extract for preventing and treating kiwifruit canker preferably includes the following components: Radix Isatidis extract.

[0008] In the present invention, the Chinese medicinal extract for preventing and treating kiwifruit canker preferably comprises the following components: Pseudo-Actinidia sylvestris extract.

[0009] In the present invention, the Chinese medicinal extract for preventing and treating kiwifruit canker preferably comprises the following components: Radix Isatidis extract and Radix Aconiti Lateralis Preparata extract.

[0010] In the present invention, the Chinese medicinal extract for preventing and treating kiwifruit canker is preferably composed of the following components: Radix Isatidis extract and Radix Aconiti Lateralis Preparata extract.

[0011] The mass ratio of the Radix Isatidis extract and the Rhizoma Cibotii extract is preferably (1-9):(1-9), more preferably (1-7):(3-9), and even more preferably 4:6.

[0012] The present invention also provides a method for preparing a Chinese medicinal extract for preventing and treating kiwifruit canker, which comprises any one of the following methods 1 to 3:

[0013] Method 1: extracting Radix Isatidis by alcohol extraction to obtain Radix Isatidis extract;

[0014] Method 2: Using alcohol extraction to extract Pseudo-Aesculus chinensis to obtain Pseudo-Aesculus chinensis extract;

[0015] Method three: extracting Radix Isatidis by alcohol extraction to obtain Radix Isatidis extract; extracting Pseudo-Jiujie by alcohol extraction to obtain Pseudo-Jiujie extract; mixing the Radix Isatidis extract and the Pseudo-Jiujie extract to obtain.

[0016] In the present invention, the alcohol solution in the alcohol extraction method is preferably an ethanol solution.

[0017] The concentration of ethanol in the ethanol solution is preferably 50-100%, for example 75%.

[0018] In the present invention, the volume of the alcohol solution added in the alcohol extraction method is preferably 5 to 15 times, for example, 7 times, the volume of the Radix Isatidis or the Rhizoma Cibotii.

[0019] In the present invention, the number of extractions is preferably 2 to 4 times, and the time for each extraction is preferably 90 to 150 minutes.

[0020] In the present invention, the alcohol extraction method can be assisted by ultrasonic extraction, which specifically includes: mixing the Radix Isatidis or Radix Pseudoschizonepetae with an alcohol solution and placing the mixture in an ultrasonic extractor.

[0021] The ultrasonic power is preferably 400-900W.

[0022] In the present invention, after the alcohol extraction of the Radix Isatidis and / or the Rhizoma Cibotii, the steps of concentration and drying may be further included.

[0023] Wherein, the concentration method is preferably condensation reflux concentration.

[0024] The drying temperature is preferably 50-65°C.

[0025] The present invention also provides an agricultural bactericidal Chinese medicine compound prepared by a method for preparing a Chinese medicine extract for preventing and treating kiwifruit canker.

[0026] The present invention also provides a use of the aforementioned Chinese medicinal extract for preventing and treating kiwifruit canker in preventing and treating kiwifruit canker.

[0027] In the present invention, the Chinese medicinal extract for preventing and treating kiwifruit canker can be used as a therapeutic agent for kiwifruit canker pathogen Psa and / or a protective agent for kiwifruit canker pathogen Psa.

[0028] In the present invention, when the Chinese herbal medicine extract comprises a combination of a Radix Isatidis extract and a Radix Aconitum extract, the dosage concentration of the composition is preferably 25 mg / mL.

[0029] The present invention also provides an application of the isatis root extract in preventing and treating kiwifruit canker.

[0030] In the present invention, the preparation method of the Radix Isatidis extract preferably comprises the following steps: extracting the Radix Isatidis by alcohol extraction to obtain the Radix Isatidis extract.

[0031] The alcohol solution in the alcohol extraction method is preferably an ethanol solution, and the concentration of ethanol in the ethanol solution is preferably 50-100%, for example 75%.

[0032] The volume of the alcohol solution added in the alcohol extraction method is preferably 5 to 15 times, for example, 7 times, the volume of the Radix Isatidis.

[0033] The extraction is preferably performed 2 to 4 times, and the duration of each extraction is preferably 90 to 150 minutes.

[0034] The alcohol extraction method may be supplemented with ultrasonic extraction, specifically comprising: mixing the Radix Isatidis with an alcohol solution and placing the mixture in an ultrasonic extractor. The ultrasonic power is preferably 400-900W.

[0035] Wherein, after the alcohol extraction of Radix Isatidis, the method may further include the steps of concentration and drying.

[0036] The concentration method is preferably condensation reflux concentration.

[0037] The drying temperature is preferably 50-65°C.

[0038] In the present invention, the concentration of the Radix Isatidis extract is preferably not less than 0.78 mg / mL, such as 12.5 mg / mL, 25.0 mg / mL, or 50.0 mg / mL.

[0039] In the present invention, the Radix Isatidis extract can be used as a therapeutic agent for kiwifruit canker pathogen Psa and / or a protective agent for kiwifruit canker pathogen Psa.

[0040] The present invention also provides an application of the pseudo-nine knotweed extract in preventing and treating kiwifruit canker.

[0041] In the present invention, the preparation method of the Pseudo-Aesculapius extract preferably comprises the following steps: extracting the Pseudo-Aesculapius by alcohol extraction to obtain the Pseudo-Aesculapius extract.

[0042] The alcohol solution in the alcohol extraction method is preferably an ethanol solution, and the concentration of ethanol in the ethanol solution is preferably 50-100%, for example 75%.

[0043] Wherein, the added volume of the alcohol solution in the alcohol extraction method is preferably 5 to 15 times, for example, 7 times, the volume of the pseudo-nine-section.

[0044] The extraction is preferably performed 2 to 4 times, and the duration of each extraction is preferably 90 to 150 minutes.

[0045] The alcohol extraction method can be supplemented with ultrasonic extraction, specifically comprising: mixing the Pseudo-Nine-joints with an alcohol solution and placing the mixture in an ultrasonic extractor. The ultrasonic power is preferably 400-900W.

[0046] Wherein, after the alcohol extraction of the pseudo-nine knotweed, the process may further include the steps of concentration and drying.

[0047] The concentration method is preferably condensation reflux concentration.

[0048] The drying temperature is preferably 50-65°C.

[0049] In the present invention, the dosage concentration of the Pseudo-Aconiti Lateralis Preparata extract is preferably not less than 0.78 mg / mL, such as 12.5 mg / mL, 25.0 mg / mL, or 50.0 mg / mL.

[0050] In the present invention, the Pseudo-Nine-node Extract can be used as a therapeutic agent for kiwifruit canker pathogen Psa and / or a protective agent for kiwifruit canker pathogen Psa.

[0051] The positive progress effect of the present invention is:

[0052] (1) The Chinese herbal extract of the present invention has the effect of preventing and treating kiwifruit canker bacteria and has the potential to be developed into a new type of botanical fungicide.

[0053] (2) The raw materials used in the present invention are traditional Chinese medicinal materials, which have less impact on humans, animals, the environment and other factors than chemical pesticides. BRIEF DESCRIPTION OF THE DRAWINGS

[0054] Figure 1 This is the inhibition zone of Radix Isatidis extract against kiwifruit canker pathogen at different concentrations.

[0055] Figure 2 The inhibition zones of Pseudo-nine-node extract against kiwifruit canker pathogen at different concentrations.

[0056] Figure 3 The morphological and structural changes of kiwifruit canker pathogens after treatment with Chinese herbal medicine extracts, where a: blank control group; b: Isatis indigotica MIC treatment group; c: Pseudo-nine-node MIC treatment group.

[0057] Figure 4 The figure shows the effects of Chinese herbal medicine extracts on the DNA content of kiwifruit canker pathogens, where 1, 2, and 3 are Pseudo-nine-node; 4, 5, and 6 are Radix Isatidis; 7 is CK, where CK is the DNA band of M228 pathogen; and M is Maker in the electrophoresis test.

[0058] Figure 5 These are the symptom changes of kiwifruit branches inoculated with Psa, where B: Radix Isatidis extract; J: Pseudo-Nine-joint extract; C: Kasugamycin; +: inoculate the fungus first and then add the drug; -: add the drug first and then inoculate the fungus. DETAILED DESCRIPTION

[0059] The present invention is further illustrated by way of examples below, but the present invention is not limited to the scope of the examples. Experimental methods in the following examples where specific conditions are not specified were performed according to conventional methods and conditions, or selected according to the product specifications.

[0060] The Radix Isatidis used in the following Examples and Comparative Examples was obtained from the dried root of Isatis tinctoria L., a Cruciferae plant, and the Psychotria tutcheri Dunn was obtained from the dried branches and leaves of Psychotria tutcheri Dunn, a Rubiaceae plant.

[0061] The reagents and raw materials used in the following examples are all commercially available.

[0062] Example 1

[0063] Preparation of Radix Isatidis Extract: Take Radix Isatidis powder in a conical flask, add 75% ethanol solution until it is immersed, the added volume of ethanol solution is 7 times the volume of the medicine, place it in an ultrasonic cleaning machine for ultrasonication for 90-150 minutes, filter the Chinese medicine ethanol mixture after ultrasonication to obtain the first extraction filtrate, then repeat the extraction of the Chinese medicine residue according to the above method twice, mix the three extraction filtrates, place it in a rotary evaporator, heat it in a 55°C water bath, condense and reflux to concentrate, and then place it in an oven at 50-65°C to dry to obtain an extract, and obtain Radix Isatidis extract with an extraction rate of 31.97%.

[0064] Preparation of Pseudo-nine-section extract: Take the Pseudo-nine-section Chinese medicine powder in a conical flask, add 75% ethanol solution until it is immersed, the added volume of the ethanol solution is 7 times the volume of the medicine, place it in an ultrasonic cleaning machine for ultrasonication for 90-150 minutes, filter the Chinese medicine ethanol mixture after ultrasonication to obtain the first extraction filtrate, then repeat the extraction of the Chinese medicine residue according to the above method twice, mix the three extraction filtrates, place it in a rotary evaporator, heat it in a 55°C water bath, condense and reflux to concentrate, and then place it in an oven at 50-65°C to dry to obtain an extract, and obtain the Pseudo-nine-section extract with an extraction rate of 21.64%.

[0065] Effect Example 1

[0066] This example describes the inhibitory effect of Chinese herbal extracts on the growth of kiwifruit canker bacteria.

[0067] The strong pathogenic bacteria of kiwifruit canker (Pseudomonas syringae pv.actinidiae M228) were used as the test pathogens to test the antibacterial effects of the Radix Isatidis extract and the Pseudomonas syringae extract in Example 1 at different concentrations.

[0068] Use a pipette to take 20 μL of the bacterial solution of kiwifruit canker M228 stored at -80°C, spread it on the LB plate culture medium in the clean bench, activate it at 25°C for 48 hours and wait for colonies to form. Then pick the activated bacteria and streak them on the LB plate culture medium and culture them at 25°C for 48 hours. After colonies are formed, select the colonies and inoculate them into liquid LB culture medium and shake them at 28°C and 200rpm for 12 hours. Centrifuge the culture obtained by shaking at 8000rpm for 1 minute, take the precipitate, rinse it with sterile water 3 times and shake it well, and measure the concentration of the bacterial solution with a spectrophotometer. Dilute the bacterial solution with sterile water to OD 600 =1, the concentration is about 1×10 8 For a bacterial suspension with a CFU / mL, pipette 2 mL of the suspension and mix with LB medium onto a Petri dish. After solidification, punch a hole in the plate with a 5 mm borer. Remove the punched medium and add 10 μL of the agent. Incubate the plate at 28°C for 2 days and observe the size of the inhibition zone. Measure the diameter of the inhibition zone using the cross-hatch method and calculate the inhibition rate using the formula: Inhibition rate (%) = [(inhibition zone diameter - 5) / inhibition zone diameter)] × 100.

[0069] Effects of Radix Isatidis Extract and Radix Pseudoschizonepetae Extract on the inhibition zone size of M228 Figure 1 、 Figure 2 As shown in Table 1, Table 2 shows the inhibition rates of Radix Isatidis extract and Pseudo-Nine Section extract on kiwifruit canker pathogens.

[0070] Table 1 Inhibition zone size of Chinese herbal medicine extracts against kiwifruit canker pathogens at different concentrations

[0071]

[0072]

[0073] Table 2 Inhibition rate of Chinese herbal medicine extracts on kiwifruit canker pathogens

[0074]

[0075] It can be seen that the diameter of the inhibition zone of the Radix Isatidis extract and the Pseudo-Nine Section Extract at a concentration of 25.0 mg / mL is greater than 10.0 mm; and the inhibition rate against kiwifruit canker pathogens at a concentration of 100 mg / mL is greater than 60%.

[0076] Effect Example 2

[0077] This example describes the determination of the minimum inhibitory concentration of Chinese herbal extracts against Actinidia ulcerans.

[0078] The MIC value of the biological reagent to be tested was determined by the double dilution method. Seven test tubes numbered 1 to 7 were taken and 2 mL of sterile water was added. The tubes were sterilized in a sterilizing pot for use. 2 mL of the test solution with a concentration of 2.5 mg / mL was added to test tube No. 1. After shaking, 2 mL of the mixed solution was added to test tube No. 2. The operation was repeated until tube No. 6. The 2 mL of solution in tube No. 6 was discarded so that the volume of the solution in each test tube was the same. Tube No. 7 was set as the blank control. In addition, three groups of repeated treatments were set for each pathogen. The concentrations of Isatis root extract and Pseudo-nine-section extract in test tubes 1 to 7 are shown in Table 3. Finally, 2 mL of the diluted pathogen suspension with a bacterial count of 106 CFU / mL was added to test tubes No. 1 to 7 respectively, and placed in an incubator at a suitable temperature for culture. After the bacterial solution in the blank control test tube became turbid, the OD of each test tube was measured using a spectrophotometer. 600 Values, compared, measured OD 600 The lowest drug concentration that is closest to the blank control without bacterial solution is the minimum inhibitory concentration.

[0079] The results are shown in Table 3. When the concentration of Radix Isatidis extract is 0.78 mg / mL, its OD 600 The value is closest to the blank control without bacterial solution, and the inhibitory effect is the smallest. As the concentration increases, the lower the OD value, the more significant the antibacterial effect. Therefore, the minimum inhibitory concentration (MIC) of Radix Isatidis extract is 0.78 mg / mL. 600 The value is 0.78 mg / mL. As the concentration increases, the OD value decreases, so its minimum inhibitory concentration (MIC) is 0.78 mg / mL.

[0080] Table 3 Effects of Chinese herbal medicine extracts at different concentrations on bacterial OD 600 Impact of value

[0081]

[0082] Effect Example 3

[0083] This example describes the scanning electron microscopic observation of the effects of Chinese herbal extracts on kiwifruit canker bacteria.

[0084] 1.5 mL of cultured bacterial suspension was placed in a 2 mL sterile centrifuge tube and centrifuged at 7000 rpm at 4°C for 1 minute. Each treatment was repeated three times. The centrifuged bacteria were resuspended and washed with 1 mL of PBS solution, centrifuged again, and the culture medium was washed away. The bacteria were resuspended in 1.5 mL of PBS solution and various amounts of the traditional Chinese medicine mother solution were added to achieve a minimum inhibitory concentration (MIC) (i.e., 0.78 mg / mL). The centrifuge tubes were then placed in a shaking incubator and incubated at 28°C and 180 rpm for 10 hours. After the ethyl allicin treatment was complete, the samples were removed and centrifuged at 7000 rpm at 4°C for 1 minute. The samples were then resuspended in 1 mL of PBS solution and centrifuged again, repeated three times, and the drug solution was washed away. The bacteria from the three parallel centrifuge tubes were combined into one centrifuge tube. Finally, 1 mL of 2.5% glutaraldehyde fixative was added to each tube and fixed at 4°C for 10 hours. After the sample reacts with glutaraldehyde, it is centrifuged at 7000 rpm at 4°C for 1 minute. The sample is then resuspended in 1 mL of PBS and centrifuged again three times to remove the 2.5% glutaraldehyde. The sample is then replaced with 1 mL of 30%, 50%, 70%, 90%, and 100% ethanol, sequentially, for 10 minutes each replacement. The sample is diluted and dripped onto a 6mm coverslip and allowed to air-dry. Conductive adhesive is applied to the sample stage, and the coverslip with the sample is placed on top. The sample is then sprayed with gold, and the sample is placed on the scanning electron microscope stage for observation and photography.

[0085] from Figure 3 As can be seen, the bacteria in the blank control group (a) were rod-shaped, normal in morphology, plump, and with intact cell walls and membranes. At a concentration of 0.78 mg / mL, the bacteria in the Isatis Radix MIC-treated group (b) were severely wrinkled, with damaged outer walls and broken into short segments. The bacteria in the Pseudo-Nine-Segment MIC-treated group (c) adhered to each other, exhibiting significant distortion and even defects in some areas, leading to leakage of bacterial contents. This suggests that the Chinese herbal extract can significantly inhibit the pathogenic bacteria of kiwifruit canker.

[0086] Effect Example 4

[0087] This example describes the effect of Chinese herbal extracts on the DNA content of Actinidia ulcerans.

[0088] DNA contains the genetic information necessary for the synthesis of RNA and proteins and is an essential macromolecule for the development and normal functioning of organisms. A reduction in and damage to DNA content prevents normal gene expression, hindering the synthesis of intracellular enzymes and receptors and ultimately leading to cell death. The brightness of DNA bands in agarose gel retardation electrophoresis is proportional to the size of DNA molecules. The brightness of the bands can reflect the DNA content, so the difference in DNA content between the treated and control groups can be explained by comparing the brightness of the DNA bands. MIC and MBC concentrations of traditional Chinese medicine extracts and sterile water were added dropwise to bacterial cultures in the logarithmic growth phase, with three replicates for each treatment. DNA from the two treatment groups was extracted according to the instructions of the bacterial DNA extraction kit, and then separated, detected, stained, and imaged on a 1% agarose gel, followed by photographing and analysis.

[0089] from Figure 4 It can be seen that the DNA bands in the blank control group are clearer than those in the treatment group, indicating that at a concentration of 0.78 mg / mL, after treatment with Isatis Radix and Pseudo-Nine-section Extracts, the DNA content of kiwifruit canker pathogens is less than that of normal bacteria. At the same time, the DNA bands in the Pseudo-Nine-section Treatment Group are clearer than those in the Isatis Radix Treatment Group, which also proves that at the MIC concentration, the Pseudo-Nine-section Extract has a better bactericidal effect and a lower bacterial DNA content. In summary, after treatment with Isatis Radix and Pseudo-Nine-section Extracts, the cell permeability of the bacteria changes, causing the DNA molecules inside the cells to leak out, thereby reducing the content of DNA molecules in the cells.

[0090] Effect Example 5

[0091] This example describes the protective and therapeutic effects of Chinese herbal extracts on detached kiwifruit branches.

[0092] (1) Branch collection: Select one-year-old healthy and uniformly growing lignified branches with a diameter of about 0.8 cm, cut them into small branches of 10 cm each, and seal both ends of the branches with paraffin to reduce water loss. Set 3-6 replicates for each sample.

[0093] (2) Branch treatment: Each branch was surface-disinfected with 0.6% sodium hypochlorite, rinsed twice with sterile water, and air-dried. Subsequently, a sterile blade was used to incise the branches 1.5 cm from the branch endpoint. Two incisions were made on each branch, 2 mm wide and deep to the phloem.

[0094] a. Therapeutic effects of the agent on branches: Inoculate with diluted Psa bacterial solution, dropping 10 μL of the solution onto each incision. Incubate the branches in a 16°C incubator for 24 hours to allow the pathogen to establish a parasitic relationship with the host. Then, add a medium-inhibitory concentration of the traditional Chinese medicine compound to the incision. The treatment with sterile water served as a negative control, and the treatment with kasugamycin served as a positive control.

[0095] b. Protective effect of the agent on the branches: add a medium concentration of Chinese herbal compound to the incision, use the treatment with sterile water as the negative control, and the treatment with kasugamycin as the positive control, place the branches in a constant temperature box for 24 hours and then inoculate the bacterial solution.

[0096] (3) Branch culture: After the bacterial solution penetrates into the branch wound, place the branch with the wound facing upward on a tray with wet filter paper on the bottom. Cover the branch with a film to keep it moist. Keep the humidity in the tray at around 80%. Place it in an artificial climate box at 22-24°C and a relative humidity of 80% for 12 hours of light culture and 12 hours of dark culture.

[0097] (4) Lesion measurement: After 20 days of incubation, the branch cortex was removed with a sterile knife and the length of the brown lesions caused by the pathogen on the branch was measured. The preventive effect was calculated according to the formula: preventive effect (%) = [(control lesion length - treatment lesion length) / control lesion length] × 100.

[0098] Figure 5 Shown are the symptom changes of kiwifruit branches inoculated with Psa.

[0099] Table 4 Lesion length of kiwifruit branches after inoculation with Psa (therapeutic effect)

[0100]

[0101] Table 5 Lesion length of kiwifruit branches after inoculation with Psa (protective effect)

[0102]

[0103] In the test of in vitro branch inoculation to determine the therapeutic effect of the agent, the results are shown in Table 4, among which the positive control kasugamycin had an inhibition rate of 57.1% at a concentration of 25.0 mg / mL. In comparison, at this concentration, the isatis root extract showed a better therapeutic effect, with an average inhibition rate of 42.9%, indicating that the isatis root extract can effectively prevent the further spread of kiwifruit canker bacteria and play a therapeutic role, while the average inhibition rate of the pseudo-nine-section extract at a concentration of 25.0 mg / mL was 21.4%.

[0104] In the test of inoculating detached branches to determine the protective effect of the agents, it can be seen from Table 5 that at a concentration of 25.0 mg / mL, the average inhibition rate of kasugamycin was 52.5%, and the average inhibition rates of isatis root and pseudo-nine knot extracts also reached 41.2% and 47.1%, respectively, showing good protective effects. This indicates that isatis root and pseudo-nine knot extracts can resist the invasion of kiwifruit canker pathogens through their protective effects.

[0105] Example 2

[0106] The Radix Isatidis extract and the Psoralea corylifolia extract prepared in Example 1 were mixed in different percentages by mass to obtain an agricultural bactericidal Chinese medicine compound, wherein the mass ratios of the Radix Isatidis extract and the Psoralea corylifolia extract were 100:0, 90:10, 80:20, 70:30, 60:40, 50:50, 40:60, 30:70, 20:80, 10:90, and 0:100, respectively.

[0107] Effect Example 6

[0108] This example describes the inhibitory effect of the agricultural bactericidal Chinese medicine compound of Example 2 on the plate growth of kiwifruit canker bacteria.

[0109] The diameters of the inhibition zones of the agricultural bactericidal Chinese medicine compound at different mass ratios were calculated according to the method in Example 1. The concentration of the agricultural bactericidal Chinese medicine compound was 25.0 mg / mL. Table 6 shows the diameters of the inhibition zones of the Radix Isatidis extract and the Pseudo-Jiujie extract at different mass ratios.

[0110] Table 6 Inhibition zone diameters of Radix Isatidis and Pseudo-Cypripedium wilfordii extracts at different ratios

[0111]

[0112] Note: a, b, c represent significant differences among different treatments.

[0113] As can be seen from Table 6, the synergistic effect of the Radix Isatidis extract and the Pseudo-Nine-joints extract in the range of 70:30 to 10:90 was significantly different compared with the single application (P<0.05), and at the composite ratio of 40:60, the antibacterial effect on the kiwifruit canker pathogen M228 was the best, with the diameter of the inhibition zone reaching 15.7 mm, which was 5.4 mm larger than the diameter of the inhibition zone of the Radix Isatidis extract applied alone, and 4.6 mm larger than the diameter of the inhibition zone of the Pseudo-Nine-joints extract applied alone. Therefore, the combination of the Radix Isatidis extract and the Pseudo-Nine-joints extract of the present invention produced a synergistic effect.

Claims

1. A Chinese medicinal extract for preventing and treating kiwifruit canker, characterized in that: The active ingredient of the Chinese medicine extract is Pseudo-Aconiti Lateralis Preparata extract; The preparation method of the pseudo-nine joint extract includes: extracting the pseudo-nine joint by alcohol extraction, concentrating and drying to obtain the pseudo-nine joint extract; the alcohol solution in the alcohol extraction method is an ethanol solution, the added volume of the alcohol solution in the alcohol extraction method is 5 to 15 times the volume of the pseudo-nine joint, the number of extractions is 2 to 4, and the time of each extraction is 90-150 minutes; the alcohol extraction method is assisted by ultrasonic extraction, specifically including: mixing the pseudo-nine joint with the alcohol solution and placing it in an ultrasonic extractor.

2. The Chinese medicinal extract for preventing and treating kiwifruit canker according to claim 1, wherein It meets one or more of the following conditions: ① The concentration of ethanol in the ethanol solution is 50-100%; ②The ultrasonic power is 400-900W; ③ The concentration method is condensation reflux concentration; and ④ the drying temperature is 50-65°C.

3. A Chinese medicinal extract for preventing and treating kiwifruit canker, characterized in that: The effective ingredients of the traditional Chinese medicine extract are the following components: Radix Isatidis extract and Radix Aconitum extract, and the mass ratio of the Radix Isatidis extract to the Radix Aconitum extract is (1-7):(3-9); The preparation method thereof comprises: The method comprises extracting the isatis root by alcohol extraction, concentrating and drying, and obtaining the isatis root extract; the alcohol solution in the alcohol extraction method is an ethanol solution, the volume of the alcohol solution added in the alcohol extraction method is 5 to 15 times the volume of the isatis root, the number of extractions is 2 to 4, and the time of each extraction is 90 to 150 minutes; and assisting in ultrasonic extraction in the alcohol extraction method, specifically comprising: mixing the isatis root with the alcohol solution and placing the mixture in an ultrasonic extractor; The method comprises extracting the pseudo-nine knotweed by alcohol extraction, concentrating and drying, and obtaining a pseudo-nine knotweed extract; the alcohol solution in the alcohol extraction method is an ethanol solution, the added volume of the alcohol solution in the alcohol extraction method is 5 to 15 times the volume of the pseudo-nine knotweed, the number of extractions is 2 to 4, and the time of each extraction is 90 to 150 minutes; and the alcohol extraction method is assisted by ultrasonic extraction, specifically comprising: mixing the pseudo-nine knotweed with the alcohol solution and placing the mixture in an ultrasonic extractor; The Radix Isatidis extract and the Rhizoma Cibotii extract are mixed to obtain the product.

4. The Chinese medicinal extract for preventing and treating kiwifruit canker according to claim 3, wherein It meets one or more of the following conditions: ① The concentration of ethanol in the ethanol solution is 50-100%; ②The ultrasonic power is 400-900W; ③ The concentration method is condensation reflux concentration; ④ The drying temperature is 50-65°C; and ⑤ the mass ratio of the Radix Isatidis extract to the Pseudo-Cyperus rotundus extract is 4:

6.

5. A method for preparing a Chinese medicinal extract for preventing and treating kiwifruit canker, characterized in that: It adopts method one or method two: Method 1: The method comprises extracting the pseudo-nine knotweed by alcohol extraction, concentrating and drying, and obtaining a pseudo-nine knotweed extract; the alcohol solution in the alcohol extraction method is an ethanol solution, the added volume of the alcohol solution in the alcohol extraction method is 5 to 15 times the volume of the pseudo-nine knotweed, the number of extractions is 2 to 4, and the time of each extraction is 90 to 150 minutes; and the alcohol extraction method is assisted by ultrasonic extraction, specifically comprising: mixing the pseudo-nine knotweed with the alcohol solution and placing the mixture in an ultrasonic extractor; Method 2: The method comprises extracting the isatis root by alcohol extraction, concentrating and drying, and obtaining the isatis root extract; the alcohol solution in the alcohol extraction method is an ethanol solution, the volume of the alcohol solution added in the alcohol extraction method is 5 to 15 times the volume of the isatis root, the number of extractions is 2 to 4, and the time of each extraction is 90 to 150 minutes; and assisting in ultrasonic extraction in the alcohol extraction method, specifically comprising: mixing the isatis root with the alcohol solution and placing the mixture in an ultrasonic extractor; The method comprises extracting the pseudo-nine knotweed by alcohol extraction, concentrating and drying, and obtaining a pseudo-nine knotweed extract; the alcohol solution in the alcohol extraction method is an ethanol solution, the added volume of the alcohol solution in the alcohol extraction method is 5 to 15 times the volume of the pseudo-nine knotweed, the number of extractions is 2 to 4, and the time of each extraction is 90 to 150 minutes; and the alcohol extraction method is assisted by ultrasonic extraction, specifically comprising: mixing the pseudo-nine knotweed with the alcohol solution and placing the mixture in an ultrasonic extractor; The Radix Isatidis extract and the Rhizoma Cibotii extract are mixed in a mass ratio of (1-7):(3-9) to obtain the product.

6. The method for preparing the Chinese medicinal extract for preventing and treating kiwifruit canker according to claim 5, wherein: It meets one or more of the following conditions: ① The concentration of ethanol in the ethanol solution is 50-100%; ②The ultrasonic power is 400-900W; ③ The concentration method is condensation reflux concentration; ④ The drying temperature is 50-65°C; and ⑤ the mass ratio of the Radix Isatidis extract to the Pseudo-Cyperus rotundus extract is 4:

6.

7. Application of an isatis root extract in preventing and treating kiwifruit canker; The preparation method of the Radix Isatidis extract comprises the following steps: The isatis root is extracted by alcohol extraction, and after concentration and drying, the isatis root extract is obtained; The alcohol solution in the alcohol extraction method adopts ethanol solution, the added volume of the alcohol solution in the alcohol extraction method is 5 to 15 times the volume of the isatis root, the number of extractions is 2 to 4 times, and the time of each extraction is 90-150 minutes. The alcohol extraction method is assisted by ultrasonic extraction, specifically comprising: mixing the isatis root with the alcohol solution and placing it in an ultrasonic extractor.

8. The use according to claim 7, characterized in that It meets one or more of the following conditions: ① The concentration of ethanol in the ethanol solution is 50-100%; ②The ultrasonic power is 400-900W; ③ The concentration method is condensation reflux concentration; and ④ the drying temperature is 50-65°C.

9. Application of an extract of Actinidia chinensis in preventing and treating kiwifruit canker; The preparation method of the Pseudo-Aconiti Lateralis Preparata extract comprises: The euphorbia pulex was extracted by alcohol extraction, and after concentration and drying, the euphorbia pulex extract was obtained; The alcohol solution in the alcohol extraction method adopts ethanol solution, the added volume of the alcohol solution in the alcohol extraction method is 5 to 15 times the volume of the pseudo-nine joints, the number of extractions is 2 to 4 times, and the time of each extraction is 90-150 minutes. The alcohol extraction method is assisted by ultrasonic extraction, specifically comprising: mixing the pseudo-nine joints with the alcohol solution and placing them in an ultrasonic extractor.

10. The use according to claim 9, characterized in that It meets one or more of the following conditions: ① The concentration of ethanol in the ethanol solution is 50-100%; ②The ultrasonic power is 400-900W; ③ The concentration method is condensation reflux concentration; and ④ the drying temperature is 50-65°C.

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